The long-term goal of this multi-disciplinary project is to understand how epithelial stem cells in the skin establish distinct patterns of gene activation and silencing during their differentiation into specialized cell lineages and how these genetic programs are re-organized during skin regeneration and aging. Recent data revealed that in addition to signaling/transcription factor-dependent regulatory mechanisms, lineage-specific gene expression programs are also regulated epigenetically, i.e., via modulation of covalent DNA/histone modifications, as well as through higher-order chromatin remodeling and establishment of long- range associations or interactomes between the genes and their enhancer elements in 3D nuclear space. In normal differentiating cells, lineage-specific long-range chromatin interactions provide structural frameworks for cell-specific transcription or silencing. Importantly, these interactions are substantially re- organized during cell transition towards malignancy, while genes located closely in topologically associated chromatin domains frequently serve as sites for chromosomal translocations in cancers. Our recent studies revealed that during skin development, transcription factor-dependent and epigenetic regulatory mechanisms are intimately linked to each other via p63 transcription factor, which plays a novel, previously unrecognized role in regulation of expression of chromatin remodeling genes Satb1 and Brg1. In this proposal, we will further address a fundamental biological problem on how epigenetic machinery operates in concert with p63 transcription master regulator to control gene expression in skin epithelial stem cells during their differentiation in specialized (epidermal, har follicle) cell lineages. In particular, we will elucidate how higher-order chromatin remodeling associated with gene activation and silencing is controlled and how functional interactomes between the genes and their enhancer elements or other genes are formed in the keratinocytes during terminal differentiation. These questions will be addressed via two Specific Aims: 1. Define a role of p63 and its target genes Brg1 and Satb1 in the control of higher-order chromatin remodeling and topological interactomes of the lineage-specific genes and their enhancer elements in skin epithelial stem cells and their progenies. 2. Identify the role of p63 and its target Polycomb Cbx4 gene in the control of formation of the repressive chromatin compartments to silence non-keratinocyte lineage genes and selected cell cycle- associated genes in epithelial stem cells and their progenies. This project will have a fundamental impact on our current knowledge of epigenetic mechanisms that regulate genome reorganization in stem cells during their differentiation in the skin and will promote the progress towards the development of novel epigenetic drugs as new paradigm for treatment of skin disorders.